Age-related effects of fenofibrate on the hepatic expression of sirtuin 1, sirtuin 3, and lipid metabolism-related genes.

BACKGROUND: Sirtuin 1 (Sirt1) and sirtuin 3 (Sirt3) participate in the regulation of lipid metabolism. Our aim was to investigate the effects of the hypolipemic drug fenofibrate (FN) on hepatic Sirt1 and Sirt3 expression, in relation to the expression of lipid metabolism-related genes and in the context of aging. METHODS AND RESULTS: Young and old male Wistar rats were fed standard chow or supplemented with 0.1% or 0.5% FN for 30 days (n=7-10 in each group). In young rats, 0.1% FN did not affect Sirt1 expression, however, 0.5% FN decreased Sirt1 and both doses reduced Sirt3 protein levels. In old rats, 0.5% FN decreased hepatic Sirt1 mRNA and both doses reduced Sirt1 protein levels, but not Sirt3 expression. Although hepatic Pparα protein levels did not change, FN treatment of young rats induced Cpt1b expression, whereas Lcad, Acox1, Pmp70, and Hmgcs2 expression increased only after 0.1% FN, and Fas2 expression decreased after 0.5% FN. In the liver of old rats, both doses increased Cpt1b and Lcad expression. Only 0.1% FN increased Pmp70 and Hmgcs2 expression, and only 0.5% FN increased Acox1 and Fas2 mRNA levels. CONCLUSIONS: Treatment with fenofibrate at low or high doses may downregulate the expression of Sirt1 and Sirt3 proteins in the rat liver. The dosage of FN affects molecular changes, and aging alters the response to 0.5% FN.

Sestrins' Expression in the Liver Is Not Altered by Short-Term Calorie Restriction in Young and Old Rats.

INTRODUCTION: Short-term calorie restriction (SCR) may have a positive impact on health. We hypothesized that sestrins, a family of stress-inducible proteins (Sesn1, Sesn2, Sesn3) are involved in the response to SCR in the liver. METHODS: Young-adult (4-month) and old (24-month) male Wistar rats were either fed ad libitum (control groups) or received 60% of food intake on a daily basis for 30 days (SCR groups). In blood sera, biochemical parameters and fibroblast growth factor 21 (FGF21) concentration were measured (ELISA). Liver samples were collected for analyses of genes' expression (real-time PCR) and protein levels (Western blotting). RESULTS: SCR caused improvements in blood glucose and lipids and parameters of liver function but did not affect the serum FGF21 concentration. SCR caused changes typically associated with calorie restriction in the gene expression of fatty acid synthase (fasn), ATP-citrate lyase (acly), and sirtuin 1 (sirt1). In the liver of young SCR rats, protein level of Sesn2 tended to increase, while Sesn3 tended to decrease, accompanied by reduced sesn3 expression. In old SCR rats, reduced sesn1 expression was reflected by decreasing trend for Sesn1 content. Activation of AMP-activated protein kinase (phospho-Thr172) and protein content of peroxisome proliferator-activated receptor gamma coactivator 1-alpha were not affected by SCR. CONCLUSION: Sestrins' hepatic expression is only minimally affected by SCR in young and old rats. We propose that sestrins may not be a major effector of mild SCR in the liver of young or old rats.

Short-term fenofibrate treatment improves ultrastructure of hepatocytes of old rats.

INTRODUCTION: Fenofibrate (FN) is a hypolipemic drug used for the treatment of mixed dyslipidemia. Since in our previous study FN administration to young and old rats adversely affected the serum activity of liver marker enzymes, we decided to examine the effects of FN on liver ultrastructure of young and old animals. MATERIAL AND METHODS: Young and old rats were fed standard rodent chow supplemented with 0.1% FN for 30 days. Liver samples obtained from animals under full anesthesia were processed by routine methods to obtain ultrathin and histological sections for the examination by light microscopy (LM) and transmission electron microscopy (TEM). Furthermore, liver lysates were analyzed by Western blotting for the expression of the autophagy-related proteins LC3A/B and beclin 1. RESULTS: The ultrastructure of hepatocytes in both age groups was well-preserved, with the presence of abundant mitochondria, numerous peroxisomes and lysosomes, glycogen stored in the form of rosettes, and occasionally autolysosomes. However, hepatocytes of old control rats contained less mitochondria and peroxisomes, and more lipid droplets than cells of young animals. The effects of FN on liver ultrastructure were age-depended. FN increased the relative number of mitochondria and peroxisomes in the hepatocytes of old, and did not affect their number in young rats. Moreover, FN decreased and increased the relative number of lipid droplets in the hepatocytes of old and young rats, respectively. At the LM level, Oil Red O staining revealed smaller and larger lipid droplets within hepatocytes and non-parenchymal liver cells. In the livers of young and old rats lipid droplets were distributed mainly in the periportal zones of hepatic lobules. Morphometric analysis confirmed that livers of control old rats contained more lipid-stainable areas than those of young ones; however, no effect of FN was observed either in young or old rats. Despite larger size of autolysosomes and autophagic vacuoles in hepatocytes of old rats, the expression of autophagy-related proteins did not differ in the livers of control and fenofibrate-treated young and old animals. CONCLUSIONS: The results of our study suggest that fenofibrate, apart from its hypolipemic action, may have beneficial effect on the energy metabolism in the liver of old individuals by increasing the number of mitochondria and peroxisomes in hepatocytes.

Regulation of mitochondrial dynamics in 2-methoxyestradiol-mediated osteosarcoma cell death.

Osteosarcoma (OS) is one of the most malignant tumors of childhood and adolescence. Research on mitochondrial dynamics (fusion/fission) and biogenesis has received much attention in last few years, as they are crucial for death of cancer cells. Specifically, it was shown that increased expression of the cytoplasmic dynamin-related protein 1 (Drp1) triggers mitochondrial fission (division), which activates BAX and downstream intrinsic apoptosis, effectively inhibiting OS growth. In the presented study, human OS cells (metastatic 143B OS cell line) were incubated with 2-methoxyestradiol (2-ME) at both physiologically and pharmacologically relevant concentrations. Cell viability was determined by the MTT assay. Confocal microscopy and western blot methods were applied to examine changes in Drp1 and BAX protein levels. Mitochondrial Division Inhibitor 1, MDIVI-1, was used in the study to further examine the role of Drp1 in 2-ME-mediated mechanism of action. To determine quantitative and qualitative changes in mitochondria, electron microscopy was used. 2-ME at all used concentrations increased mitochondrial fission and induced autophagy in OS cells. At the concentration of 1 µM 2-ME increased the area density of mitochondria in OS cells. Subsequent, upregulated expression of Drp1 and BAX proteins by 2-ME strongly suggests the activation of the intrinsic apoptosis pathway. We further observed 2-ME-mediated regulation of glycolytic state of OS cells. Therefore, we suggest that changes of mitochondrial dynamics may represent a novel mechanism of anticancer action of 2-ME. This finding may open new approaches to improve the efficacy of chemotherapy in the treatment of OS, however, it has to be confirmed by in vivo studies.

Fenofibrate impairs liver function and structure more pronounced in old than young rats.

INTRODUCTION: Since old animals are known to accumulate lipids in some organs, we compared effects of fenofibrate (FN) on systemic lipid metabolism, activity of liver marker enzymes and structure in young and old rats. MATERIAL AND METHODS: Young and old rats were fed chow supplemented with 0.1 % or 0.5 % FN. After 30 days, intraperitoneal glucose tolerance test (IPGTT) was performed, and blood and liver samples were collected. RESULTS: In young rats, 0.1 % FN, but not 0.5 % FN, decreased serum Chol by 74 %, and did not affect TG levels at either doses. In old rats, 0.5 % FN, but not 0.1 % FN, decreased Chol and TG level by 56 % and 49 %, respectively. In young rats, 0.1 % and 0.5 % FN increased serum activity of ALP by 227 % and 260 %, respectively, and did not affect AST and ALT activities. In old rats, only 0.5 % FN increased serum ALP activity by 150 %, respectively. In old rats, neither dose of FN affected serum AST activity, and only 0.5 % FN increased serum ALT activity by 200 %. The histological examination of liver structure revealed that both doses of FN impaired lobular architecture, expansion of bile canaliculi, and degeneration of parenchymal cells with the presence of cells containing fat droplets; administration of FN increased area occupied by collagen fibers. CONCLUSIONS: Although 0.5 % FN decreased serum Chol concentration, it increased serum ALP activity and impaired liver structure in both in both age groups of rats. Thus, FN treatment should be under the control of liver function, especially in older patients.